This article details a protocol for isolating and perfusing the heart of an anesthetized mouse using a Langendorff apparatus. The procedure involves careful dissection and cannulation to ensure successful perfusion for downstream applications.
To begin, place an anesthetized mouse in the supine position on a dissection board. Make an incision to access the thoracic cavity and expose the heart. Locate the aorta and its branches - the left common carotid artery and brachiocephalic artery. Transect these arteries to isolate the heart.
Immediately, immerse the excised heart into a Petri dish containing a nutrient-rich oxygenated buffer solution. Concomitantly, insert a cannula into the heart such that its tip reaches the ascending aorta. Use a suture to ligate the cannula. Connect the cannula to a syringe containing oxygenated perfusion solution.
Inject the perfusion solution into the heart. This step allows the blood to leave the coronary arteries and effuse from the pulmonary veins. Next, connect the cannulated heart to the Langendorff apparatus. Use peristaltic pump to perform perfusion at a constant flow rate.
Under these conditions, the buffer perfuses into the heart. This creates cannulation pressure that closes the aortic valves, causing the buffer to move through the coronary arteries. Eventually, the buffer perfuses the heart muscles retrogradely and later effuses through the pulmonary veins. A successfully perfused heart is now ready for further downstream applications.
After euthanizing an 8- to 10-week-old male C57BL/6 mouse, use tissue forceps to lift the skin of the xiphoid. Then, using tissue scissors, make a minor lateral incision through the skin. Perform a blunt dissection between the skin and fascia. Extend the skin incision toward the axillae in a V-shape on both sides and continue the incision through the rib cage.
Then, using tissue forceps, clamp the sternum and deflect the rib cage upward to fully expose the heart and lungs. Using curved forceps, peel off the pericardium. If the thymus gland covers the great vessels, use two curved forceps to tear the thymus gland toward both sides. Then, gently pull the base of the heart toward the tail until the aorta and its branch arteries are visible as a Y-shaped blood vessel.
Transect the aorta at the left common carotid artery. Then, cut the brachiocephalic artery. Excise the heart and immediately immerse it in a Petri dish containing Tyrode's solution to wash and pump out the residual blood. Then, transfer the heart to another Petri dish containing solution 1.
Under a stereomicroscope, using fine iris scissors, trim any surplus tissue. Expel air bubbles from the syringe. Then, with the assistance of two straight tying forceps, perform retrograde aortic cannulation, taking care that the whole cannulation process is performed under the liquid surface. Adjust the cannulation depth such that the cannula tip is in the ascending aorta, taking care not to penetrate the aortic valves.
Then, with a pre-knot 3-0 suture, ligate the aorta to the cannula notch. Gently inject solution 1 from the syringe to flush out the residual blood. Then, connect the cannulated heart to the Langendorff apparatus, taking care not to introduce any air bubbles into the heart. After connecting the cannulated heart to the Langendorff apparatus, perfuse the heart with solution 1 for approximately 2 minutes.
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